LAIiORATORY AND GREENHOUSE STUDIES. 131 



These tubes were allowed to stand over night, and on the followmg 

 day dilutions of the tubes were made in the customary manner and 

 plates were poured, using Dolt's synthetic medium. 



In two days every one of the 10 plates showed pink colonies, 

 and where there were more than two or three colonies the agar was 

 entirely reddened. In some plates the colonics were few, while in 

 others they were numerous. For the most part they were round 

 and to all appearances like Bacillus coli. The fact that they red- 

 dened the litmus and grew well in this lactose medium is good evidence 

 of their identity. 



Transfers were made on May 21 from the pink colonies to litmus 

 milk and incubated at 37° C. In 24 hours the four tubes were red, 

 coagulated, and showed abundant whev. Transfers from these 

 litmus-milk tubes were made to nitrate bouillon on May 28 and three 

 days afterwards were tested for the reduction of nitrates to nitrites. 

 Each one of the tubes showed the reduction well. 



On June 2 transfers were made from the litmus-milk tubes to 

 fermentation tubes containing peptone and neutral red to test for the 

 canary-yellow color. The tubes were incubated at 37° C. After 

 two days light clouding took place, but no gas formation nor reduc- 

 tion of color. This behavior being entirely contrary to that of 

 Bacillus coli, the medium was tested for dextrose, which it should 

 have for the complete reaction. It was found by the use of Feliling's 

 solution that not a particle of reducing sugar was in the fermentation 

 tubes, so a fresh medium containing the peptone, dextrose, and 

 neutral red was made up. These tubes were incubated at 37° C. 

 After 18 hours a small amount of gas appeared, but little change in 

 color. In 24 hours the tubes showed 31 to 34 millimeters of gas, 

 and each one was changed to the canary yellow in the closed end 

 and to a bright red in the open end. 



Transfers from each of these four fermentation tubes were made 

 to beef gelatin and incubated for 48 hours at 37° C. At the end of 

 that time all the tubes showed heavy precipitate, several large clots 

 in suspension, and moderate films. The tubes were placed in an 

 ice box and allowed to harden. Eventually each one of these became 

 solidified and showed absolutely no sign of liquefaction of the gelatin. 



Thus, in this third experiment the same organism that was inocu- 

 lated, viz, Bacillus coli, was isolated from the decayed tissues. 



Experiment No. 4. 



On May 7 Miss McCulloch inoculated several coconuts under the 

 supervision of the writer. Strains of Bacillus coli (Hitchings and 

 VI-ll-V-09) which had not been in the hands of the writer at all 

 were used. The coconuts for this experiment had but recently been 



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